This invention relates to the treatment of autoimmune diseases and transplantation rejection in mammals. More specifically, the present invention relates to the use of a narrowly-defined group of methimazole derivatives and tautomeric cyclic thiones for the purposes described herein.
A primary function of immune response in mammals is to discriminate self from non-self antigens and to eliminate the latter. The immune response involves complex cell to cell interactions and depends primarily on three major types of immune cells: thymus derived (T) lymphocytes, bone marrow derived (B) lymphocytes, and macrophages. Immune response is mediated by molecules encoded by the major histocompatibility complex (MHC). The two principal classes of MHC molecules, Class I and Class II, each comprise a set of cell surface glycoproteins (see Stites, D. P. and Terr, A. I. (eds), xe2x80x9cBasic and Clinical Immunologyxe2x80x9d, Appelton and Lange, Norwalk, Conn./San Mateo, Calif., 1991). MHC Class I molecules are found on virtually all somatic cell types, although at different levels in different cell types. By contrast, MHC Class II molecules are normally expressed only on a few cell types, such as lymphocytes, macrophages and dendritic cells.
Antigens are presented to the immune system by antigen presenting cells in the context of Class I or Class II cell surface molecules, for example, CD4+ helper T-lymphocytes recognize antigens in association with Class II MHC molecules, and CD8+ cytotoxic lymphocytes (CTL) recognize antigens in association with Class I gene products. It is currently believed that MHC Class I molecules function primarily as the targets of the cellular immune response, while Class II molecules regulate both the humoral and cellular immune response (Klein, J. and Gutze, E., xe2x80x9cMajor Histocompatibility Complexxe2x80x9d, Springer Verlag, New York, 1977; Unanue, E. R., Ann. Rev. Immunology, 2:295-428, (1984)). MHC Class I and Class II molecules have been the focus of much study with respect to research in autoimmune diseases because of their roles as mediators or initiators of immune response. MHC Class II antigens have been the primary focus of research in the etiology of autoimmune diseases, whereas MHC Class I antigens have historically been the focus of research in transplantation rejection.
Numerous experimental animal models for human disease have linked aberrant expression and/or function of MHC Class I and MHC Class II antigens to the autoimmune disease process, for example, insulin-dependent diabetes mellitus (IDDM) (Tisch and McDevitt, Cell 85: 291-297 (1996)), systemic lupus erythematosus (SLE) (Kotzin, Cell 85: 303-306 (1996)), and uveoretinitis (Prendergast et al., Invest. Opthalmol. Vis. Sci. 39: 754-762 (1998)).
The pathological link between MHC Class I and/or Class II expression and disease has been examined in many of these model systems using a variety of biochemical and genetic approaches. However, the strongest evidence for aberrant MHC gene function as a mediator of autoimmune disease stems from transgenic animal models in which the MHC genes have been inactivated. Using MHC Class I deficient animals resistance to the autoimmune disease processxe2x80x94and hence the dependence of autoimmunity upon MHC gene expressionxe2x80x94can be directly demonstrated in animal models for IDDM (Serreze et al., Diabetes 43: 505-509 (1994)), and SLE (Mozes et al., Science 261: 91-93 (1993)).
Moreover, the dependence of the progressive multifocal inflammatory autoimmune disease phenotype exhibited by TGF-betal deficient transgenic mice (Shull et al., Nature 359: 693-699 (1992); Kulkarni et al., Proc. Natl. Acad. Sci. 90: 770-774 (1993); Boivin et al., Am. J. Pathol. 146: 276-288 (1995)) on MHC Class II expression has recently been demonstrated using MHC Class II deficient animals. Specifically, TGF-betal deficient animals lacking MHC Class II expression are without evidence of inflammatory infiltrates, circulating antibodies, or glomerular immune complex deposits (Letterio et al., J. Clin. Invest. 98: 2109-2119 (1996)).
In addition to the information supportive of MHC Class I and Class II antigens as critical for the development of autoimmunity in animal models there is equally strong evidence linking autoimmune processes with expression of MHC Class I and MCH Class II antigens in humans.
Graves"" disease is a relatively common autoimmune disorder of the thyroid. In Graves"" disease, autoantibodies against thyroid antigens, particularly the thyrotropin receptor (TSHR), alter thyroid function and result in hyperthyroidism (Stites, D. P. and Terr, A. I. (eds), xe2x80x9cBasic and Clinical Immunologyxe2x80x9d, Appleton and Lang, Norwalk, Conn./San Mateo, Calif., 1991, pp. 469-470)). Thyrocytes from patients with Graves"" disease have aberrant MHC Class II expression and elevated MHC Class I expression (Hanafusa et al., Lancet 2:1111-1115 (1983); Bottazzo et al., Lancet 2:1115-1119 (1983); Kohn, et al., in xe2x80x9cInternational Reviews of Immunology,xe2x80x9d Vol. 912, pp. 135-165, (1992)). Aberrant expression of MHC Class II and TSHR on fibroblasts, but not either alone, has recently been shown to induce Graves"" disease in mice, i.e., aberrant expression of Class II on target tissue can yield autoimmune disease in animals with normal immune systems. Thionamide therapy has historically been used to treat Graves"" disease. The most commonly used thionamides are methimazole, carbimazole and propylthiouracil. These thionamides contain a thiourea group; the most potent are thioureylenes (W. L. Green, in Werner and Ingbar""s xe2x80x9cThe Thyroidxe2x80x9d: A Fundamental Clinical Text, 6th Edition, L. Braverman and R. Utiger (eds), J. B. Lippincott Co., 1991, p. 324). The basis for thionamide therapy has, however, not focused on immune suppression. Rather, the basis has been suppression of thyroid hormone formation. Experiments suggesting an effect on immune cells, to inhibit antigen presentation or antibody formation, are largely discounted as nonphysiologic in vitro artifacts of high MMI concentration. MMI activity under those circumstances is suggested to be based on free-radical scavenger activity. See D. S. Cooper, in Werner E. Ingbar""s xe2x80x9cThe Thyroidxe2x80x9d, op. cit., pp. 712-734.
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that, like Graves"" disease, has a relatively high rate of occurrence. SLE affects predominantly women, the incidence being 1 in 700 among women between the ages of 20 and 60 (Abbus, A. K., Lichtman, A. H., Pober, J. S. (eds), xe2x80x9cCellular and Molecular Immunologyxe2x80x9d, W.B. Saunders Company, Philadelphia, 1991, pp. 360-370). SLE is characterized by the formation of a variety of autoantibodies and by 20 multiple organ system involvement (Stites and Terr, ibid, pp. 438-443). Current therapies for treating SLE involve the use of corticosteroids and cytotoxic drugs, such as cyclophosphamide. Immunosuppressive drugs, such as cyclosporin, FK506 or rapamycin suppress the immune system by reducing T cell numbers and function (Morris, P. J., Curr. Opin. in Immun., 3:748-751 (1991)). While these immunosuppressive therapies alleviate the symptoms of SLE and other autoimmune diseases, they have numerous severe side effects. In fact, extended therapy with these agents may cause greater morbidity than the underlying disease. A link between MHC Class I expression and SLE in animal models has been established. Thus, Class I deficient mice do not develop SLE in the 16/6 ID model (Mozes, et al., Science 261: 91-93 (1993)).
Women suffering from SLE who have breast cancer face particular difficulties. These individuals are immunosuppressed as a result of corticosteroid and cytotoxic drug treatment for SLE; radiation therapy for the treatment of the cancer, a current treatment of choice, would additionally exacerbate the immunosuppressed state. Further, radiation therapy can exacerbate disease expression or induce severe radiation complications. For these individuals, alternative therapies that would allow for simultaneous treatment of SLE and cancer are greatly needed.
Diabetes Mellitus is a disease characterized by relative or absolute insulin deficiency and relative or absolute glucagon excess (Foster, D. W., Diabetes Mellitus. In Stanbury, J. B., et al., The Metabolic Basis of Inherited Disease. Ch. 4, pp 99-117, 1960). Type I diabetes appears to require a permissive genetic background and environmental factors. Islet cell antibodies are common in the first months of the disease. They probably arise in part to xcex2 cell injury with leakage cell antigens but also represent a primary autoimmune disease. The preeminent metabolic abnormality in Type I diabetes is hyperglycemia and glucosuria. Late complications of diabetes are numerous and include increased atherosclerosis with attendant stroke and heart complications, kidney disease and failure, and neuropathy which can be totally debilitating. The link to HLA antigens has been known since 1970. Certain HLA alleles are associated with increased frequency of disease, others with decreased frequency. Increased MHC class I and aberrant MHC class II expression in islet cells has been described (Bottazzo et al., NEJM 313: 353-360 (1985); Foulis and Farquharson, Diabetes 35: 1215-1224 (1986)). A definitive link to MHC class I has been made in a genetic animal model of the disease. Thus MHC class I deficiency results in resistance to the development of diabetes in the NOD mouse (Sereze et al., Diabetes 43: 505-509 (1994); Wicker et al., Diabetes 43: 500-504 (1994)).
A wealth of genetic, biochemical and animal model data support a contributory role of inflammatory cytokines (e.g., IL-12, IL-18; and particularly IFN-gamma) in the autoimmune process (Sarvetnick, J Clin Invest 99: 371-372 (1997)). Studies using non-obese diabetic (NOD) mice, which spontaneously develop auto-immune diabetes reminiscent of Type I human IDDM, are particularly illustrative in demonstrating how IFN-gamma stimulated processes play critical roles in the development of autoimmunity; and how the actions of other pro-inflammatory cytokines are channeled through IFN-gamma stimulated processesxe2x80x94among which are the enhanced expression of MHC Class I and MHC Class II antigens.
IL-12 and IL-18 (IFN-gamma inducing factor) are known to act synergistically in stimulating production of IFN-gamma in T cells (Micallef et al., Eur. J. Immunol. 26: 1647-1651 (1996)). In diabetic NOD mice the systemic expression of IL-18 (Rothe et al., J. Autoimmun. 10: 251-256 (1997)) and islet expression of IL-12 are increased (Rabinovitch et al., J. Autoimmun. 9: 645-651 (1996)). Moreover, additional IL-12 accelerates autoimmune diabetes in NOD mice (Trembleau et al., J. Exp. Med. 181: 817-821 (1995)). Genetic analysis has determined the IL-18 gene maps to a chromosomal region (Idd2) associated with a genetic susceptibility for autoimmune diabetes (Kothe et al., J. Clin. Invest. 99: 469-474 (1997)). These reports support help to define a critical role for IFN-gamma in the process of autoimmunity.
The role of IFN-gamma in the autoimmune process is further substantiated by studies where IFN-gamma""s signaling capacity was abrogated in some manner. For example, transgenic NOD mice deficient in the cellular receptor for IFN-gamma (Wang et al., Proc. Natl. Acad. Sci. 94: 13844-13849 (1997)) do not develop autoimmune diabetes. NOD mice treated with a neutralizing antibody for IFN-gamma (Debray-Sachs et al., J. Autoimmun. 4:237-248 (1991)) also do not develop autoimmune diabetes. While it is somewhat surprising that the onset of diabetes is only delayed in transgenic NOD mice deficient in IFN-gamma (Hultgren et al., Diabetes 45: 812-817 (1996)), this observation only further stresses the importance of blocking the IFN-gamma signalxe2x80x94and more importantly IFN-gamma stimulated downstream eventsxe2x80x94for the effective prevention of autoimmunity in NOD mice.
Analogous observations have been made in animal models for SLE. Soluble IFN-gamma receptor blocks disease in the NZB/NZW F1 spontaneous autoimmune disease model for SLE (Ozmen et al., Eur. J. Immunol. 25: 6-12 (1995)); uveitis, where the targeted expression of IFN-gamma increases ocular inflammation (Geiger et al., Invest. Opthalmol. Vis. Sci. 35: 2667-2681 (1994)); and autoimmune gastritis, where neutralizing IFN-gamma antibody blocks disesase (Barret et al., Eur. J. Immunol. 26: 1652-1655 (1996)). Moreover, in humans treatment with IFN-gamma has been reported to be associated with the development of an SLE-like disease (Graninger et al., J. Rheumatol. 18: 1621-1622 (1991)).
It is well recognized that xcex3-IFN increases MHC class I and class II expression in many tissues and thus is linked to the action of a coregulatory molecule, the class II transactivator (Mach et al., Ann Rev Immunol 14: 301-331 (1996); Chang et al., Immunity 4: 167-178 (1996); Steimle et al., Science 265: 106-109 (1994); Chang et al., J Exp Med 180: 1367-1374 (1994); Chin et al. Immunity 1: 687-697 (1994); Montani, V. et al., Endocrinology 139: 280-289 (1998)). It is also known that MMI can inhibit IFN-increased class I and class II expression in thyroid (Saji et al., J. Clin. Endocrinology. Metab. 75: 871-878 (1992); Montani et al., Endocrinology. 139: 290-302 (1998)). Finally, it has been shown that MMI decreases expression of CIITA increased class II expression and this appears to be related to the action of MMI to enhance Y box protein gene expression; the Y box protein suppresses class II gene expression (Montani et al., Endocrinology 139: 280-289 (1998)).
As is true for autoimmune diseases, there is a great need for new and different ways of treating or preventing transplantation rejection. Transplantation rejection occurs as a result of histoincompatibility between the host and the donor; it is the major obstacle in successful transplantation of tissues. Current treatment for transplantation rejection, as for autoimmune disease, involves the use of a variety of immunosuppressant drugs and corticosteroid treatment.
Kjellin and Sandstrom, Acta Chemica Scandinavica, 23: 2879-2887 and 2888-2899 (1969), discloses a series of tautomeric cyclic thiones, i.e., oxazoline-, thiazoline-, and imidazoline-2-(3)-thiones, having methyl and phenyl groups in the 4 and 5 positions. The compounds were used for a study of thione-thiol equilibria. No pharmaceutical, or any other utility, is disclosed or suggested for these compounds.
U.S. Pat. No. 3,641,049, Sandstrom et al., issued Feb. 8, 1972, discloses N,Nxe2x80x2-dialkyl4-phenylimidazoline-2-thiones, particularly 1,3-dimethyl4-phenylimidazoline-2-thione, for use as an antidepressant agent. The dimethyl compound is also said to exhibit antiviral properties against herpes simplex and vaccinia viruses.
U.S. Pat. No. Re. 24,505, Rimington et al., reissued Jul. 22, 1958, discloses a group of imidazole compounds useful as anti-thyroid compounds.
U.S. Pat. No. 3,505,350, Doebel et al., issued Apr. 7, 1970, discloses a group of substituted 2-mercaptoimidazole derivatives which are said to be effective as anti-inflammatory agents. Illustrative compounds include 1-(4-fluorophenyl)-5-methyl-2-mercaptoimidazole and 1-methyl-5-phenyl-2-mercaptoimidazole.
U.S. Pat. No. 3,390,150, Henry, issued Jun. 25, 1968, is representative of a group of patents which disclose nitroimidazole derivatives which possess antischistosomal and antitrichomonal activity.
U.S. Pat. No. 5,051,441, Matsumoto et al., issued Sep. 24, 1991, discloses diphenyl imidazoline derivatives which are said to act as immunomodulators, showing efficiency in the treatment of rheumatoid arthritis, multiple sclerosis, systemic lupus, and rheumatic fever.
U.S. Pat. No. 4,073,905, Kummer, et al., issued Feb. 14, 1978, discloses 2-amino4-phenyl-2-imidazolines, which are said to be useful for treating hypertension.
U.S. Pat. No. 5,202,312, Matsumoto et al., issued Apr. 13, 1993, discloses imidazoline-containing peptides which are said to have immunomodulatory activity.
PCT Application WO 92/04033, Faustman, et al., identifies a method for inhibiting rejection of transplanted tissue in a recipient animal by modifying, eliminating, or masking the antigens present on the surface of the transplanted tissue. Specifically, this application suggests modifying, masking or eliminating human leukocyte antigen (HLA) Class I antigens. The preferred masking or modifying drugs are F(ab)xe2x80x2 fragments of antibodies directed against HLA-Class I antigens. However, the effectiveness of such a therapy will be limited by the hosts"" immune response to the antibody serving as the masking or modifying agent. In addition, in organ transplantation, this treatment would not affect all of the cells because of the perfusion limitations of the masking antibodies. Faustman, et al. contends that fragments or whole viruses can be transfected into donor cells, prior to transplantation into the host, to suppress HLA Class I expression. However, use of whole or fragments of virus presents potential complications to the recipient of such transplanted tissue since some viruses, SV40 in particular, can increase Class I expression (Singer and Maguire, Crit. Rev. Immunol., 10:235-237 (1991), see particularly Table 2).
British Patent 592,453, Durant, et al., identifies isothiourea compositions that may be useful in the treatment of autoimmune diseases in host versus graft (HVG) disease and assays for assessing the immunosuppressive capabilities of these compounds. However, this patent does not describe methimazole or the suppression of MHC Class I molecules in the treatment of autoimmune diseases. No tautomeric cyclic thiones are disclosed or discussed.
Several autoimmune diseases have been treated with methimazole with potential success. In one study, MMI was deemed as good as cyclosporin in treating juvenile diabetes (W. Waldhausl, et al., Akt. Endokrin. Stoffw. 8:119 (1987), and psoriasis has also been treated with MMI.
U.S. Pat. No. 5,556,754, Singer, et al. (which is equivalent to PCT Application WO 94/28897), issued Sep. 17, 1996, describes a method for treating autoimmune diseases using methimazole, methimazole derivatives and methinazole analogs. The terms xe2x80x9cmethimazole derivativexe2x80x9d and xe2x80x9cmethimazole analogxe2x80x9d are not defined or exemplified anywhere in the patent.
U.S. Pat. No. 5,310,742, Elias, issued May 10, 1994, describes the use of thioureylene compounds to treat psoriasis and autoimmune diseases. Propylthiouracil, methimazole, and thiabendazole are the only specific compounds disclosed in the patent. Examples show the use of methimazole to treat psoriasis in humans and the use of thioureylene to treat rheumatoid arthritis, lupus and transplant rejection. No methimazole analogs or derivatives are disclosed or discussed. No tautomeric cyclic thiones are disclosed or discussed.
U.S. Pat. No. 4,148,885, Renoux, et al., issued Apr. 10, 1979, describes the use of specific low molecular weight sulfur-containing compounds as immunostimulants. Methimazole, thioguanine and thiouracil are among the compounds specified. No methimazole analogs or derivatives are disclosed or discussed. No tautomeric cyclic thiones are disclosed or discussed.
U.S. Pat. No. 5,010,092, Elfarra, issued Apr. 23, 1991, describes a method of reducing the nephrotoxicity of certain drugs via the coadministration of methimazole or carbimazole (which is taught to be the pro-drug of methimazole) together with the nephrotoxic drug. No methimazole analogs or derivatives are discussed in this patent. No tautomeric cyclic thiones are disclosed or discussed.
U.S. Pat. No. 5,578,645, Askanazi, et al., issued Nov. 26, 1996, describes a method for minimizing the side effects associated with traditional analgesics. This is accomplished via the administration of a mixture of specific branched amino acids together with the analgesic compound. Methimazole is disclosed, in the background section of this patent, as a non-steroidal anti-inflammatory drug which may provide some of the side effects which this invention is said to address. No tautomeric cyclic thiones are disclosed or discussed.
U.S. Pat. No. 5,587,369, Daynes, et al., issued Dec. 24, 1996, describes a method for preventing or reducing ischemia following injury. This is accomplished by introducing dehydroepiandrosterone (DHEA), DHEA derivatives or DHEA congeners to a patient as soon as possible after the injury. The background section of this patent teaches that methimazole is a thromboxane inhibitor which has been shown to prevent vascular changes in burn wounds.
The U.S.P. Dictionary (US Pharmacopeia, Rockville, Md., 1996) includes methimazole (CAS-60-56-0) and describes it as a thyroid inhibitor.
Methimazole, therefore, is known in the art for a variety of pharmaceutical utilities: for the treatment of psoriasis (Elias), as an immunostimulant (Renoux, et al.), for the reduction of nephrotoxicity of certain drugs (Elfarra), for the minimization of side effects found with certain analgesics (Oskinasi, et al), as a thyroid inhibitor (USP Dictionary), and as a thromboxane inhibitor (Daynes, et al.). It is also taught in the Singer, et al. patent as being useful in the treatment of autoimmune diseases, such as rheumatoid arthritis and systemic lupus. While the Singer, et al. patent contains general references to the use of methimazole analogs and derivatives for these therapeutic purposes, no definition of these compounds is given and no specific compounds are suggested. The pharmacological properties of tautomeric cyclic thiones are not discussed nor related to those of methimazole derivatives.
It has now been found that a specific class of methimazole derivatives and tautomeric cyclic thiones are effective in treating autoimmune diseases and suppressing the rejection of transplanted organs, and that these compounds show clear and unexpected benefits over the use of methimazole itself. In particular, these compounds: (a) are more effective in inhibiting basal and IFN-induced Class I RNA expression and in inhibiting IFN-induced Class II RNA expression than methimazole; (b) inhibit the action of IFN by acting on the CIITA/Y-box regulatory system; (c) may be significantly more soluble than methimazole, leading to significant formulation flexibility and advantages; (d) have less adverse effects on thyroid function than methimazole; (e) have an enhanced ability to bind to targets affected by MMI; and (f) exhibit therapeutic activities in vivo. These properties are unexpected based on the known properties of methimazole and particularly the tautomeric cyclic thiones.
Finally, the present invention relates to the method by which these agents inhibit interferon-gamma actions, specifically those related to increase MHC Class I and MHC Class II expression and mediation of pro-inflammatory processes, and more specifically those processes related to the induction of autoimmune disease and/or transplant rejection.
The present invention relates to pharmaceutical compositions comprising a safe and effective amount of an active compound selected from 
wherein Y is H, C1-C4 alkyl, C1-C4 substituted alkyl, xe2x80x94NO2, or the phenyl moiety 
wherein no more than one Y group in said active compound may be the phenyl moiety; R1 is selected from H, xe2x80x94OH, halogens (F, Cl, Br or I), C1-C4 alkyl, C1-C4 substituted alkyl, C1-C4 ester or C1-C4 substituted ester; R2 is selected from H, C1-C4 alkyl or C1-C4 substituted alkyl; R3 is selected from H, C1-C4 alkyl, C1-C4 substituted alkyl or xe2x80x94CH2Ph (wherein Ph is phenyl); R4 is selected from H, C1-C4 alkyl or C1-C4 substituted alkyl; X is selected from S or O; Z is selected from xe2x80x94SR3, xe2x80x94OR3, S(O)R3 or C1-C4 alkyl; and wherein at least two of the R2 and R3 groups on said compound are C1-C4 alkyl when Y is not a phenyl moiety, and at least one Y is xe2x80x94NO2 when Z is alkyl; together with a pharmaceutically-acceptable carrier.
Preferred compounds for use in these pharmaceutical compositions have the forumlae 
wherein Y is selected from H and C1-C4 alkyl or C1-C4 substituted alkyl; R1 is selected from H, xe2x80x94OH, halogens (F, Cl, Br, or I), or C1-C4 alkyl, C1-C4 substituted alkyl, C1-C4 ester or C1-C4 substituted ester; R2 is selected from H or C1-C4 alkyl or C1-C4 substituted alkyl; R3 is selected from H, C1-C4 alkyl, C1-C4 substituted alkyl, or xe2x80x94CH2Ph; R4 is selected from H, C1-C4 alkyl or C1-C4 substituted alkyl; X is selected from S or O; and Z is selected from xe2x80x94SR3 or xe2x80x94OR3.
Particularly preferred compounds are those which have the formulae 
Preferred compounds also include those of the formulae: 
wherein R9 is selected from xe2x80x94OH, xe2x80x94M and MCH2COOxe2x80x94; and M is selected from F, Cl, Br and I.
The present invention also relates to the method of treating autoimmune diseases or transplantation rejection in a patient in need of such treatment by the administration of a safe and effective amount of the active compounds and pharmaceutical compositions described above.
The present invention also relates to in vivo assay methods which permit high efficiency screening of the effects of compounds on the expression of MHC Class I and Class II proteins.
Finally, the present invention relates to the method by which the compounds defined herein inhibit gamma interferon actions to increase MHC class I or class II expression. Gamma interferon has been linked to expression of immune disease.
As used herein, all ratios, fractions and percentages are xe2x80x9cby weightxe2x80x9d, unless otherwise specified.